High dose exposure to the major cat allergen, Feld 1, has been associated with a paradoxical decrease in sensitization. Many children who are exposed to high levels of cat allergen (>=20mu/g/g dust), but who are not allergic, have a serologic profile (IgG pos IgE/neg) that is distinct from allergic subjects (IgG p?s IgEP?S); this has been described as a modified Th2 response. Such "high dose" tolerance could explain the decrease in asthma among subjects living with a cat. Regulatory T cells induced by high dose natural exposure to allergens commonly associated with asthma have not been studied. Novel CD4+ T cell major epitopes ofFel d 1 chain 2 which selectively induce IL-10 and IFN-gamma (peptides 2:1and 2:2) have been implicated in tolerance induced during immunotherapy (IT) for cat. T cell reactivity to these epitopes is diminished in highly allergic patients with atopic dermatitis (AD)(mean IgE ab to cat -- 21 IU/ml) compared to patients with a modified Th2 response who express the major HLA-DR allele "0701. In the proposed studies, the relevance of regulatory cytokines (IL-10 and TGF-[3) to Fel d 1-specific CD4+ T cell responses will be investigated using cultures stimulated with overlapping peptides of Fel d 1. Subjects studied will include allergic patients with and without AD, DR7+ and DR7- modified Th2 responders, and control (IgG neg IgE neg) subjects. Additional mediators of tolerance will be sought using antibody-based protein arrays. Patients receiving high dose IT for cat will be included in order to compare systemic (IT) and inhalational (modified Th2) tolerance and T cell epitopes relevant to systemic tolerance will be studied by monthly monitoring of patients starting IT. The relevance of chain 2 epitope-specific T cells to the modified Th2 response will be studied using MHC restriction analysis, by enumerating cells using ELISPOT assay, and by studying phenotypic characteristics using flow cytometry. Effects of regulatory cytokines on epitopespecific T cell responses will be examined and tissue-specific homing of Feld 1-specific cells to the lungs (asthmatic subjects), gut (modified Th2) or skin (AD) will also be analyzed. The natural forms of chain 2 epitopes will be identified from peptide mixtures on the surface of antigen presenting cells using mass spectrometry. Finally, given the evidence that CD8+ T cells may also target chain 2 epitopes, the ability for class I MHC peptides of Fel d 1 to induceCD8+ T cell responses in vitro will be examined and epitope-specific CD8+ T cells will be analyzed by peptide-MHC tetramer staining. Defining cellular mechanisms which govern tolerance to cat is fundamental to understanding the factors that control the prevalence and severity of asthma and to the rational design of new therapies.